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NBAF-Liverpool

We operate MiSeq and HiSeq sequencers, giving great flexibility in platform across a range of applications, including RNAseq, CHiPseq, BiSulphite sequencing, whole genome resequencing and de novo sequencing. We typically run 2 x 100 bp reads, which on the HiSeq can produce up to 600 Gb/run. The MiSeq can produce around 12 million reads, which can be in the form of 2 x 250bp reads, can be used for 16S rDNA sequencing of 400 bp amplicons. We can also produce mate-pair libraries (2 – 10kb inserts) for scaffolding genome assemblies.

We have also recently benefited from a major investment in staff and equipment from HEFCE and UniLever, which allows us to strengthen the capacity we can offer to NERC researchers, particularly in the analysis of bacterial communities.

Pacific Biosciences single molecule sequencing

From early 2014, our Pacific Biosiences RSII SMRT sequencer allows reads of 10kb that can be applied to de novo genome assembly, haplotyping, the resolution of repetitive regions, alternate splicing analysis and base modification epigenetics applications. This is the only single molecule sequencer on the market and one of very few in the UK.

Next-generation Roche 454 FLX sequencing

We operate two Roche 454 instruments which both now use the latest Titanium release, incorporating bar-coding of samples up to 100-plex. Each 8h run of the instrument generates 1 million reads of 400-800 bp and we are able to run each instrument several times a week. We use this for a range of applications including de novo sequencing of prokaryote and eukaryote genomes, 16S rDNA diversity, genome re-sequencing, sequence capture onto Nimblegen arrays with targeted sequencing, metagenomics, sequencing of cDNA collections, and digital transcriptomics. We currently offer a service generating paired-end libraries of 3kb and 8kb, which will aid the construction of scaffolds. Indeed, we have achieved great success in assembling prokaryote genomes into single scaffolds.

Microarray analysis

We offer a complete service both for designing and fabricating cDNA or oligoarrays on robotic printers, but also for using the major commercial oligoarray platforms including Affymetrix, Agilent, Nimblegen and the Illumina BeadStation. We have the very latest 2µm Agilent scanning microscope capable of imaging the new very high density chips from Agilent and Nimblegen, as well as a new Maui hybridisation station. We can offer the Agilent eArray platform for the custom design of arrays for non-model species or specialised use. We have experience of this latter process based on public domain sequence data. We can also offer advice on optimal design and layout of arrays experiments including the detailed ANOVA statistical analysis as well as pipelines for the interpretation of responding gene lists using Gene Ontology, Reactome and network packages such as GeneGo and Ingenuity. Finally, we offer training for researchers wishing to develop their own capability and transfer it to their home labs.

Nanostring analysis

Our Nanostring nCounter system provide a unique means to perform absolute quantification of DNA and RNA sequences within complex samples accurately and without amplification. Applications include the quantification of expression for up to 200 genes, copy number repeat analysis and amplification-free quantification of species abundance within microbial communities.

Sequence capture

We offer both Nimblegen and Agilent sequence capture using solution -based methods. This is useful for targeted resequencing of only portions of the genome, such as exons. We can help researchers design capture experiments.

Next-generation IonTorrent sequencing

We have recently purchased an IonTorrent sequencer. These utalise semiconductor technology to allow very cheap and very fast sequencing runs. Currently they have a capacity of 10 – 100 Mb per run in 200bp reads, moving up to 1Gb in 400 bp reads.

Bioinformatics services

We offer a ‘one-stop shop’ for bioinformatic services in support of microarray and next-gen sequencing applications undertaken in the lab. This can be either as a fully collaborative interaction with users, or as a service role where we undertake specific tasks and then return the data to yourself. In handling your analysis in-house we can more easily link the analysis of data to the lab processing of your samples. We can also more effectively provide advice on experimental design, the implications of different approaches, and troubleshoot technical problems.

For next-gen sequencing experiments we offer an experienced sequence assembly and analysis service, including the production of databases and web-enabled presentations. We can also offer more extensive services for attended sequence verification and extraction, analysis and assembly/mapping, genome annotation and presentation on genome viewers. The level of processing and commitment from staff will be based on discussion with users, and joint assessment of needs.

Expression profiling can be undertaken using either microarrays or next-gen sequencing and ‘digital’ analysis. We are very experienced in experimental design and the implementation of large-scale experiments. The statistical analysis of data from different treatment groups can be undertaken using either the older reference-based designs or the newer and more cost-effective ANOVA interwoven loop principle. For this we generally use customized informatic pipelines but where appropriate we can also undertake analyses on the NBAF GeneSpring platform. Interpretation of the resulting list of differentially expressed gene lists can be undertaken using enrichment analysis and by mapping genes onto network depictions. We also have experience in identifying and validating diagnostic gene sets for use in specific applications.